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Bacterial Danger Sensing Protects Against Bacteriophage Predation
Bacterial adaptability to unfavourable conditions, in the environment or in their eukaryotic hosts, is essential for survival and proliferation. Bacteria also have to frequently contend with bacteriophages (simply termed phages) that pose a serious threat to bacterial survival. Phages are natural pr...
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| Format: | Dissertation |
| Sprache: | Englisch |
| Veröffentlicht: |
Philipps-Universität Marburg
2021
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| Online Zugang: | PDF-Volltext |
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| Zusammenfassung: | Bacterial adaptability to unfavourable conditions, in the environment or in their eukaryotic hosts, is essential for survival and proliferation. Bacteria also have to frequently contend with bacteriophages (simply termed phages) that pose a serious threat to bacterial survival. Phages are natural predators of bacteria and utilise bacterial cells as a host for replication and subsequently, release their progeny by lysing bacterial cells. Bacteria show augmented resilience against changing environmental conditions when they exist as matrix-embedded communities, termed biofilms. This matrix comprises of secreted polysaccharides and proteins that encase bacterial cells, making them more adept at surviving phage attack as compared to their planktonic counterparts. Although biofilm formation has been shown to be an advantage in bacterial survival against phage predation, the mechanism of how bacteria sense the presence of biotic stresses, such as phages, and how they initiate biofilm formation as a response is unknown.
To investigate phage-bacteria interactions, the model organism Vibrio cholerae was used. V. cholerae is a human pathogen, responsible for causing the disease cholera. V. cholerae cells form biofilms to survive in their aquatic environment, as well as in the human host. Furthermore, in both of these environments, V. cholerae cells encounter phages that have been shown to constrict the growth of V. cholerae communities and contribute to their evolution. One of the lytic phages that was co-isolated with V. cholerae, responsible for infecting a range of V. cholerae strains, called Vibriophage N4, was used as the viral agent in this study.
The results described in this thesis show that V. cholerae actively forms biofilms in response to the exposure of Vibriophage N4. This bacterial response was neither caused by the selection of phage resistant mutants, nor by hyper matrix-producing mutants. A combined approach of proteomics and transcriptomics uncovered that cells initiated the production of biofilm matrix components upon phage exposure. Biofilm matrix production was also confirmed using fluorescence confocal microscopy. When embedded in the biofilm matrix, V. cholerae cells were protected from phage predation. However, phage infection was successful at early time points prior to biofilm formation, and counterintuitively, biofilm formation was always preceded by initial cell lysis. This led to the hypothesis that phage-induced cell lysis was necessary for bacteria to elicit a biofilm response. By exposing V. cholerae cells to sonicated bacterial lysates, it was confirmed that biofilm formation in V. cholerae was triggered not by the phages themselves, but by a component of lysed bacterial cells. Moreover, this biofilm-inducing factor was found to be general to lysates obtained from various Gram-negative and -positive bacteria. By identifying the cellular fraction from which the biofilm-inducing factor originated, the signal for inducing biofilm formation was determined to be peptidoglycan. The detection of peptidoglycan fragments from lysed cells served as an indirect signal for the presence of lysis-inducing entities (such as phages), and therefore, this signalling was referred to as bacterial danger sensing. Transcriptomics was used to characterise the bacterial response to peptidoglycan and consequently, genes related to the production of an intracellular secondary messenger molecule, c-di-GMP, were found to be upregulated. Additionally, genes encoding biofilm matrix components were also upregulated, similar to the proteome and transcriptome profile of cells that survived phage infection. The production of c-di-GMP and biofilm matrix in V. cholerae cells during peptidoglycan exposure was visualised by fluorescence confocal microscopy. As c-di-GMP has been known to play a crucial role in inducing bacterial biofilm formation, it was likely that peptidoglycan exposure induced biofilm formation in V. cholerae via c-di-GMP. Peptidoglycan was also found to be a relevant signal for inducing biofilm formation in Pseudomonas aeruginosa, suggesting that danger sensing could be conserved in bacteria. These results demonstrate how danger sensing induces bacterial biofilm formation to facilitate a rapid and general response to protect cells against phages, and potentially, other lytic stresses. |
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| Umfang: | 167 Seiten |
| DOI: | 10.17192/z2021.0501 |